Arginine Silicate For Periodontal Disease

ABSTRACT

Methods for treating and/or preventing periodontal disease with arginine silicate are disclosed. Methods may include the steps of identifying an individual in need of treatment for, or in need of prevention of, periodontal disease and administering an effective amount of an arginine-silicate complex to said individual. Arginine silicate may also be used to ameliorate one or more symptoms of periodontal disease. In some aspects, arginine silicate may be used to restore gum health and/or stop or reverse recession of the gums and/or the loss of gum tissue in the oral cavity.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. patent application Ser. No.15/068,350 filed on Mar. 11, 2016, which claims priority to U.S.Provisional Patent Application No. 62/133,076, entitled “ARGININESILICATE FOR PERIODONTAL DISEASE,” filed Mar. 13, 2015, the entirecontent of each of which is hereby incorporated by reference.

BACKGROUND Field

The present disclosure is related to the treatment and/or prevention ofperiodontal disease. More specifically, disclosed herein is the use ofarginine silicate for the treatment and/or prevention of periodontaldisease and associated symptoms.

Related Art

Periodontal disease, gum disease, periodontitis, and/or pyorrhea, referto diseases affecting the periodontium and/or teeth. Periodontitisgenerally involves the progressive loss of the gum tissue and/oralveolar bone around the teeth.

SUMMARY

Some embodiments are directed to the use of arginine and silicate and/orinositol and/or complexes thereof for use in the treatment and/orprevention of periodontal disease. More particularly, some embodimentsdisclose that an arginine silicate complex may be used as dietarysupplement to promote gum health, promote the soft tissue health of theoral cavity, treat or prevent periodontal disease, including, but notlimited to, gingivitis, periodontitis, tooth decay, bone mineral densityin the oral cavity and/or enamel erosion, and the like.

Symptoms of periodontitis include, but are not limited to, swollen gums,bright red or purplish gums, gums that feel tender when touched, gumsthat pull away from your teeth (recede), teeth that appear larger and/orlonger than normal, increased space between teeth, pus around the teethand/or gums, persistent bad breath, persistent bad taste in the oralcavity, one or more loose teeth, and/or a change in the way the teethfit together during biting.

Some embodiments provide methods of promoting gum health, promoting thesoft tissue health of the oral cavity, methods of treating or preventinggingivitis, treating or preventing periodontitis, treating or preventingtooth decay, improving bone mineral density in the oral cavity, and/ortreating or preventing enamel erosion.

In some embodiments, the complex is provided to an individual in needthereof. In some embodiments, the complex is administered to anindividual in need thereof. In some embodiments, the complex is providedor administered in an amount effective to improve gum health in anindividual in need thereof. In some embodiments, the complex is providedor administered in an amount effective to promote gum health. In someembodiments, the complex is provided or administered in an amounteffective to promote the health of the soft tissue of the oral cavity.

In some embodiments, the complex is provided or administered in anamount effective to treat periodontal disease. In some embodiments, thecomplex is provided or administered in an amount effective to preventperiodontal disease. In some embodiments, the complex is provided oradministered in an amount effective to treat or prevent gingivitis. Insome embodiments, the complex is provided or administered in an amounteffective to treat or prevent periodontitis. In some embodiments, thecomplex is provided or administered in an amount effective to treat orprevent tooth decay. In some embodiments, the complex is provided oradministered in an amount effective to treat or prevent decreased bonemineral density in the oral cavity.

In some embodiments, the effective amount is between about 100 mg andabout 5,000 mg. In some embodiments, the effective amount is betweenabout 250 mg and about 2,500 mg. In some embodiments, the effectiveamount is between about 500 mg and about 1,500 mg. In some embodiments,the complex is administered orally. In some embodiments, the complex isadministered parenterally. In some embodiments, the complex isadministered topically to the oral cavity. In some aspects, the complexis administered topically to the oral cavity in the form of a cream. Insome aspects, the complex is administered topically to the oral cavityin the form of a paste (e.g., toothpaste).

Some embodiments provide methods of identifying an individual in need ofan improvement in gum health, an improvement in the health of the softtissue of the oral cavity, and/or an improvement in bone mineral densityin the oral cavity. Some embodiments provide methods of identifying anindividual in need of treatment for periodontal disease, gingivitis,periodontitis, tooth decay, and/or enamel erosion. Some embodimentsprovide methods of identifying an individual at risk for developingperiodontal disease, gingivitis, periodontitis, tooth decay, and/orenamel erosion, (i.e., individuals in need of preventing theaforementioned conditions at a greater level than the generalpopulation).

In some embodiments, the complex is administered one to three timesdaily. In some embodiments, the complex is administered on aper-kilogram basis to humans or animals of different weights. Forexample, in some embodiments, the complex is administered one to threetimes daily in an amount ranging from about 2 mg/kg of body weight to2,500 mg/kg of body weight.

Disclosed herein are methods for increasing bone density in the oralcavity, including, but not limited to alveolar bone and maxillary bone.The method may include identifying a patient having periodontal diseaseor symptoms thereof. The method may also include administering an amountof arginine silicate effective to increase bone density in the patient'soral cavity.

Methods for ameliorating one or more symptoms of periodontal diseaseand/or periodontitis are also disclosed. The method may includeidentifying a patient having periodontal disease or symptoms thereof.The method may also include administering an amount of arginine silicateeffective to ameliorate the one or more symptoms.

Methods for reducing inflammation of the gums are also disclosed. Themethod may include identifying a patient having receding gums. Themethod may also include administering an amount of arginine silicateeffective to reduce the patient's gum inflammation. The method mayinclude administering an amount of arginine silicate effective toreduce, stop, or reverse gum tissue loss and/or gum recession.

Some embodiments provide methods of treating or preventing periodontaldisease in an individual in need thereof comprising identifying anindividual in need of treatment for, or in need of prevention of,periodontal disease, and administering an effective amount of anarginine-silicate complex to said individual.

Some embodiments provide methods for ameliorating one or more symptomsof periodontal disease comprising identifying an individual havingperiodontal disease or symptoms thereof, and administering an amount ofarginine silicate effective to ameliorate the one or more symptoms.

Some embodiments provide methods of ameliorating gum tissue loss,comprising identifying an individual having receding gums; andadministering an amount of arginine silicate effective to stop orreverse gum tissue loss.

In some embodiments, periodontal disease comprises tooth decay. In someembodiments, the periodontal disease comprises enamel erosion. In someembodiments, the periodontal disease comprises gingivitis. In someembodiments, the periodontal disease comprises decreased gum health.

In some embodiments, the effective amount of an arginine-silicatecomplex is administered orally. In some embodiments, the effectiveamount of an arginine-silicate complex is topically administered to theoral cavity. In some embodiments, the effective amount of anarginine-silicate complex is administered parenterally.

In some embodiments, the identifying an individual in need of treatmentfor, or in need of prevention of, periodontal disease comprises physicalexamination by a dental hygienist, dentist, oral surgeon, orthodontist,or medical doctor.

In some embodiments, the arginine-silicate complex comprises arginine,silicate, and inositol.

BRIEF DESCRIPTION OF THE DRAWINGS

While the experiments disclosed herein involve the treatment of rats, itis to be understood that the treatment of humans and other subjects isalso contemplated. Moreover, various dosing amounts, schedules,formulations, and delivery systems are also fully contemplated.

FIG. 1A graphically depicts the Il-1B, MMP-8, RANK, RANK-L, OPG and MPOexpression levels from periodontal tissue (a—significant differencesfrom the group as control (p<0.05); b—significant differences from thegroup as periodontitis (p<0.05); c—significant differences from thegroup as ASI I (p<0.05)). No common superscript differs significantly atthe level of p<0.01 by Fisher's multiple comparison test.

FIG. 1B depicts western blot strips Il-1B, MMP-8, RANK, RANK-L, OPG andMPO expression levels from periodontal tissue.

FIG. 2 graphically depicts inflammatory cell infiltration (“ICI”) levelsfor different treatment populations.

FIG. 3 depicts images of alveolar bone (“AK”), ligament (“BD”), dental(“D”), epithelial tissue (“ED”), inflammatory cells (“EH”), pulpa (“P”),periodontal ligament (“PL”). PANEL (A) shows the control: non-ligatedtreatment; (HE×40), little visible. PANEL (B) shows the control,non-ligated treatment; (HE×100) no visible ICI. PANEL (C) shows theligature-only; (HE×40), dense ICI. PANEL (D) shows the ligature-only(HE×100), dense ICI. PANEL (E) shows the ligature plus ASI with a doseof 1.81 mg/kg of diet; (HE×40), slightly visible ICI. PANEL (F) showsthe ligature plus ASI with a dose of 1.81 mg/kg of diet; (HE×100)slightly visible ICI. PANEL (G) shows the ligature plus ASI with a doseof 3.62 mg/kg of diet; (HE×40), slightly visible ICI. PANEL (H) showsthe ligature plus ASI with dose of 3.62 mg/kg of diet; (HE×100),slightly visible ICI.

FIG. 4 graphically depicts the bone mineral density results shown inTable 4.

FIG. 5 graphically depicts the mean of bone loss parameter results shownin Table 5 (a—significant differences from the group as control(p<0.001); b—significant differences from the group as periodontitis(p<0.001); c—significant differences from the group as ASI I (p<0.001)).

FIG. 6 graphically depicts the rate of bone loss results shown in Table6 (a—significant differences from the group as control (p<0.001),b—significant differences from the group as periodontitis (p<0.001)).

FIG. 7 depicts 3D micro-computed tomography images from the controlgroup.

FIG. 8 depicts 3D micro-computed tomography images from the L group.

FIG. 9 depicts 3D micro-computed tomography images from the ASI I group.

FIG. 10 depicts 3D micro-computed tomography images from the ASI IIgroup.

DETAILED DESCRIPTION

The present disclosure relates to new uses of an arginine silicatecomplex. In general, the complex is produced by combining arginine, asilicate salt and inositol. Although the compositions described hereingenerally contain arginine, silicate and inositol, it may be referred tothroughout the specification as “arginine silicate,” “arginine silicateinositol,” “ASI,” or “complex.”

The complexes of the present disclosure may comprise and/or consistessentially of ASI for use in the treatment and/or prevention ofperiodontal disease. In some aspects, the present compositions may beused to ameliorate one or more biochemical markers of one or moredisease states associated with periodontal disease. In some aspects,methods disclosed herein include administering to a subject a dose of acomposition containing an arginine silicate inositol complex that iseffective to treat and/or ameliorate one or more systems associated withperiodontal disease

Some embodiments provide a method for ameliorating the symptomsassociated with a bone or cartilage disorder in the oral cavity of anindividual in need thereof, comprising administering to the individualan effective amount of the arginine silicate inositol complex. In someaspects, the disclosed complexes may be used to decrease inflammation ofthe gums. For example, in some aspects, arginine silicate can be used toreduce inflammatory cell infiltration in periodontal tissue. In someaspects, arginine silicate can be used to slow the rate of lacunaeresorption. In some aspects, arginine silicate can be used to decreasethe length of the distance between the mesial cemento-enamel junctionand the mesial crestal bone in patients in need thereof. In someaspects, arginine silicate can be used to decrease the length of thedistance between the roof of furcation crestal bone and the roof offurcation. Arginine silicate may also be used to promote gum growth suchthat the cementoenamel junction of a tooth is no longer exposed.

In some embodiments, an individual is a mammal. In some embodiments, themammal is a human. In some embodiments, the mammal is mouse, rat, dog,cat, or equine.

In some aspects, the disclosed complexes may be used to decrease theamounts of one or more biomarkers associated with periodontitis.

The compositions of the present disclosure may comprise and/or consistessentially of ASI for use in the treatment and/or prevention of toothdecay. In some aspects, arginine silicate can be used to increase bonedensity in the oral cavity and/or prevent the loss of bone mass in theoral cavity. In some aspects, the arginine silicate can be used toincrease tooth strength and/or enamel levels. In some aspects, thearginine silicate can be used to prevent and/or reduce the rate of boneresorption in the oral cavity.

Some embodiments may be administered parenterally, orally,intravenously, intraarterially, intramuscularly, topically, or in anyother systemic or localized fashion, in appropriate dosage units, asdesired.

The term “parenteral” used herein includes subcutaneous, intravenous,intraarterial, injection or infusion techniques, without limitation.However, oral administration is preferred, including topicaladministration to the oral cavity (e.g., as a mouthwash or mouth rinse).Some embodiments may be in a powder form, liquid form or a combinationof powder and liquid forms. For oral administration, the complexes maybe provided as a tablet, aqueous or oral suspension, dispersible powderor granule, emulsion, hard or soft capsule, syrup or elixir.

Compositions intended for oral use may be prepared according to anymethod known in the art for the manufacture of pharmaceuticalcompositions and such compositions may contain one or more of thefollowing agents: sweeteners, flavoring agents, coloring agents,preservatives, solubilizers, wetting agents, stabilizers, colorants,antioxidants, coating agents and diluents. The sweetening agents andflavoring agents will increase the palatability of the preparation.Tablets containing silicate inositol in an admixture with non-toxicpharmaceutically acceptable excipients suitable for tablet manufactureare acceptable. Such excipients include inert diluents such as calciumcarbonate, sodium carbonate, lactose, calcium phosphate or sodiumphosphate; granulating and disintegrating agents such as corn starch oralginic acid; binding agents such as starch, gelatin or acacia; andlubricating agents such as magnesium stearate, stearic acid or talc.Tablets may be uncoated or may be coated by known techniques to delaydisintegration and absorption in the gastrointestinal tract and therebyprovide a sustained action over a longer period of time. For example, atime delay material such as glyceryl monostearate or glyceryl distearatealone or with a wax may be employed.

Formulations for oral use may also be presented as hard gelatin capsuleswherein the active ingredient is mixed with an inert solid diluent, forexample calcium carbonate, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, such as peanut oil, liquid paraffin or olive oil.Formulations for oral use may also include solutions and/or suspensions.Liquid formulations may be provided. In some aspects, the formulationsmay include a mouthwash or mouth rinse.

In some aspects, arginine silicate may be added to food that is designedfor animals. For example, the formulation may be added to and/orcomprise a pet treat or biscuit, for example, a dog biscuit or a cattreat

Aqueous suspensions may contain the complexes disclosed herein inadmixture with excipients suitable for the manufacture of aqueoussuspensions. Such excipients include suspending agents, dispersing orwetting agents, one or more preservatives, one or more coloring agents,one or more flavoring agents and one or more sweetening agents such assucrose or saccharin.

Oil suspensions may be formulated by suspending the active ingredient ina vegetable oil, such as arachis oil, olive oil, sesame oil or coconutoil, or in a mineral oil such as liquid paraffin. The oil suspension maycontain a thickening agent, such as beeswax, hard paraffin or cetylalcohol. Sweetening agents, such as those set forth above, and flavoringagents may be added to provide a palatable oral preparation. Thesecompositions may be preserved by an added antioxidant such as ascorbicacid. Oil suspensions may be formulated by suspending the activeingredient as a dispersible powder or granule in water, in an admixturewith a dispersing or wetting agent, a suspending agent and one or morepreservatives. Additional excipients, for example sweetening, flavoringand coloring agents, may also be present. Syrups and elixirs may beformulated with sweetening agents such as glycerol, sorbitol or sucrose.Such formulations may also include a demulcent, a preservative, aflavoring or a coloring agent.

In some embodiments, the complexes are administered as a compositioncomprising an orally acceptable carrier in a product such as mouthwash,toothpaste, dental cream, chewing gum, denture adhesive, or a softpliable tablet (“chewie”). As used herein, an “orally acceptablecarrier” refers to a material or combination of materials that are safefor use in the compositions disclosed herein, commensurate with areasonable benefit/risk ratio.

The compositions for parenteral administration may be in the form of asterile injectable preparation, such as a sterile injectable aqueous oroleaginous suspension. This suspension may be formulated according tomethods well known in the art using suitable dispersing or wettingagents and suspending agents. The sterile injectable preparation mayalso be a sterile injectable solution or suspension in a non-toxicparenterally-acceptable diluent or solvent, such as a solution in1,3-butanediol. Suitable diluents include, for example, water, Ringer'ssolution and isotonic sodium chloride solution. In addition, sterilefixed oils may be employed conventionally as a solvent or suspendingmedium. For this purpose, any bland fixed oil may be employed includingsynthetic mono- or diglycerides. In addition, fatty acids such as oleicacid may likewise be used in injectable preparations.

The disclosed complexes can also be administered by inhalation. In thisadministration route, an arginine silicate inositol complex can bedissolved in water or some other pharmaceutically acceptable carrierliquid for inhalation, or provided as a dry powder, and then introducedinto a gas or powder that is then inhaled by the patient in anappropriate volume so as to provide that patient with a measured amountof an arginine silicate inositol complex.

Controlled release vehicles are well known to those of skill in thepharmaceutical sciences. The technology and products in this art arevariably referred to as controlled release, sustained release, prolongedaction, depot, repository, delayed action, retarded release and timedrelease; the words “controlled release” as used herein is intended toincorporate each of the foregoing technologies.

Numerous controlled release vehicles are known, including biodegradableor bioerodable polymers such as polylactic acid, polyglycolic acid, andregenerated collagen. Known controlled release drug delivery devicesinclude creams, lotions, tablets, capsules, gels, microspheres,liposomes, ocular inserts, minipumps, and other infusion devices such aspumps and syringes. Implantable or injectable polymer matrices, andtransdermal formulations, from which active ingredients are slowlyreleased, are also well known and can be used in the disclosed methods.

Controlled release preparations can be achieved by the use of polymersto form complexes with or absorb the arginine silicate inositol complex.The controlled delivery can be exercised by selecting appropriatemacromolecules such as polyesters, polyamino acids,polyvinylpyrrolidone, ethylenevinyl acetate, methylcellulose,carboxymethylcellulose, and protamine sulfate, and the concentration ofthese macromolecule as well as the methods of incorporation are selectedin order to control release of active complex.

Controlled release of active complexes can be taken to mean any of theextended release dosage forms. The following terms may be considered tobe substantially equivalent to controlled release, for the purposes ofthe present disclosure: continuous release, controlled release, delayedrelease, depot, gradual release, long term release, programmed release,prolonged release, programmed release, proportionate release, protractedrelease, repository, retard, slow release, spaced release, sustainedrelease, time coat, time release, delayed action, extended action,layered time action, long acting, prolonged action, sustained actionmedications and extended release, release in terms of pH level in thegut and intestine, breakdown of the molecule and based on the absorptionand bioavailability.

Hydrogels, wherein an arginine silicate inositol complex is dissolved inan aqueous constituent to gradually release over time, can be preparedby copolymerization of hydrophilic mono-olefinic monomers such asethylene glycol methacrylate. Matrix devices, wherein an argininesilicate inositol complex is dispersed in a matrix of carrier material,can be used. The carrier can be porous, non-porous, solid, semi-solid,permeable or impermeable. Alternatively, a device comprising a centralreservoir of an arginine silicate inositol complex surrounded by a ratecontrolling membrane can be used to control the release of the complex.Rate controlling membranes include ethylene-vinyl acetate copolymer orbutylene terephthalate/polytetramethylene ether terephthalate. Use ofsilicon rubber depots are also contemplated.

Controlled release oral formulations are also well known. In oneembodiment, the active complex is incorporated into a soluble orerodible matrix, such as a pill or a lozenge. In another example, theoral formulations can be a liquid used for sublingual administration.These liquid compositions can also be in the form a gel or a paste.Hydrophilic gums, such as hydroxymethylcellulose, are commonly used. Alubricating agent such as magnesium stearate, stearic acid, or calciumstearate can be used to aid in the tableting process. In a preferredembodiment, transdermal patches, steady state reservoirs sandwichedbetween an impervious backing and a membrane face, and transdermalformulations, can also be used to deliver an arginine silicate inositolcomplex. Transdermal administration systems are well known in the art.One type of transdermal patch is a polymer matrix in which the activeagent is dissolved in a polymer matrix through which the activeingredient diffuses to the skin.

The terminology used in the description presented herein is not intendedto be interpreted in any limited or restrictive manner, simply becauseit is being utilized in conjunction with a detailed description ofcertain specific embodiments described herein. Furthermore, embodimentsdescribed herein can include several novel features, no single one ofwhich is solely responsible for its desirable attributes or which isessential to practicing the embodiments described herein.

The term “treating” or “treatment” does not necessarily mean total cure.Any alleviation of any undesired signs or symptoms of the disease to anyextent or the slowing down of the progress, or even prevention of thedisease or condition can be considered treatment.

As used herein, a composition that “substantially” comprises a complexmeans that the composition contains more than about 80% by weight, morepreferably more than about 90% by weight, even more preferably more thanabout 95% by weight, and most preferably more than about 97% by weightof the complex.

Throughout the specification there are references, for example, toidentifying a subject in need of administration of a ASI or in need oftreatment for periodontal disease and the like or in need of preventionof periodontal disease and the like. The term identification is notintended to be limiting and includes in each instance a belief by thesubject that the composition will benefit the subject,self-identification, and identification by third party using varioustechniques. The identification may include, but is not limited to, theassociation or identification with one or more conditions selected fromthe group consisting of: periodontal disease, gingivitis, gum disease,tooth decay, gum recession, periodontitis, enlarged gum pockets, swollengums, bleeding gums, loose teeth, sensitive teeth, persistent badbreath, and/or poor oral hygiene. Identification may include physicalexamination by a dental hygienist, dentist, oral surgeon, orthodontist,or medical doctor. In some aspects, individuals may self-identify.

The amount of a complex that will be effective in the treatment of aparticular disorder or condition disclosed herein will depend on thenature of the disorder or condition, and can be determined by standardclinical techniques. In addition, in vitro or in vivo assays mayoptionally be employed to help identify optimal dosage ranges.

The precise dose to be employed in the compositions will also depend onthe route of administration, and the seriousness of the disease ordisorder, and should be decided according to the judgment of thepractitioner and each circumstances. However, suitable dosage ranges fororal administration are generally about 0.001 milligram to 5000milligrams of a total ASI per kilogram body weight. In preferredembodiments, the oral dose is 0.01 milligram total ASI complex to 1000milligrams per kilogram body weight, more preferably 0.1 milligram to100 milligrams per kilogram body weight, more preferably 0.5 milligramto 25 milligrams per kilogram body weight, and yet more preferably 1milligram to 10 milligrams per kilogram body weight. Oral compositionspreferably contain 10% to 95% active ingredient.

Advantageously, an effective amount may be between about 2 mg and about2,500 mg. More advantageously, the effective amount is between about 500mg and about 1,000 mg. For the average 70 kg man, this may equate to adosage of between about 3.6 and 14 mg/kg (250-2,500 mg) and betweenabout 7.1 mg/kg and 14 mg/kg (500 mg-1,000 mg), respectively. Effectivedoses may be extrapolated from dose-response curves derived from invitro or animal model test systems. Such animal models and systems arewell known in the art.

The compositions disclosed herein can preferably be formulated withother active ingredients. For example, compositions disclosed herein maybe formulated in combination with fluoride. The transitional phrase“consisting essentially of” limits the scope of a claim to the specifiedmaterials or steps and those that do not materially affect the basic andnovel characteristics of the claimed invention. Thus, for example, acomposition consisting essentially of arginine silicate would notinclude other ingredients that are known to treat and/or preventperiodontal disease (e.g. fluoride).

The complexes may be administered once or twice a day. In some aspects,the complexes are administered three times a day. For example, thecomplexes may be administered before, after, or during a meal. In someaspects the complexes are administered before, during, or after toothbrushing.

EXAMPLES Example 1

All animals and surgical procedures were handled in accordance withguidelines of the Chancellor's Animal Research Committee of the Officefor Protection of Research Subjects at the University of Inonu, Malatya,Turkey (2013/A-12). Fifty-two (2-month-old) female Sprague-Dawley ratsweighing 138 (Experimental Research Center of Inonu University) werekept in temperature-controlled cages (approximately 25° C.), exposed toa 24-hr light-dark cycle of equal time, and had free access to water andfood ad libitum.

The fifty-two rats were randomly divided into four groups: thenon-ligated treatment (Control) group (n=13), the ligature-only (L)group (n=13), the ligature plus ASI (“arginine-silicate-inositol;”Nutrition 21, NY, USA, see, e.g. U.S. Pat. Nos. 5,707,970 and 7,576,132)with dose of 1.81 mg/kg of diet group (designated “ASI I”), (n=13), theligature plus ASI with dose of 3.62 mg/kg of diet group (designated “ASIII”) (n=13).

For all the studies described herein, the well-characterized ligaturemodel of placing a sterile wire ligature around the crown of the rightfirst maxillary molar was utilized. Rats were anesthetized with Rompun®,10 mg/kg ve Xylazine®, 40 mg/kg ketamin, and a sterile 28-gauge wireligature was placed around the cervical portion of the right firstmolar. Animals continued to receive basal diet or basal dietsupplemented with 1.81 or 3.62 g ASI/kg diet (Proctor et al., 2005) for8 weeks before ligature placement.

Animals were monitored weekly to ensure presence of the ligature, andthe ligature was adjusted if necessary. At the end of the experiment,animals were euthanized, blood was collected via cardiac puncture, andserum biomarkers including Ca, Mg, phosphorus, alkaline phosphatase, andCRP (C-reactive protein) were measured. Whole maxillas were removed,placed in 10% formalin for 48 hours, and stored in 70% ethanol.

A histologic evaluation was performed by a single examiner (I.O.) whowas masked to the identity of the samples. The specimens were fixed in a10% neutral-buffered formalin solution and demineralized in an aqueous10% formic acid solution. The specimens were then dehydrated, embeddedin paraffin, and sectioned along the molars in a mesiodistal plane forhematoxylin and eosin staining, as described by Toker et al. Lightmicroscopy (Nikon, Tokyo, Japan), and assessment was performed on thetwo sections with a thickness of about 6 mm, corresponding to the buccaland lingual areas between the first and second molars where ligatureshad been placed.

The areas of alveolar bone and interdental septum were analyzed underlight microscopy, considering parameters including inflammatory cellinfiltration (“ICI”) of the periodontal tissues, existing resorptionlacunae (osteoclast surfaces), osteoblastic activity (forming surfaces)and the number of osteoclasts. ICI was determined by semiquantitativescoring as not visible ICI (score=0), slightly visible ICI (score=1),and dense ICI (score=2). Osteoclasts were counted based on theirmorphology.

Bones were imaged by micro-computed tomographic (μCT) scanning(SkyScan1172 Compact MKT (Kontich, Belgium)) at 16-μm resolution, andvolumetric data were converted to DICOM format and imported in the Recon1.6.9.4 SkyScan (Contich, Belgium) to generate 3D and multiplanarreconstructed images. An oral and maxillofacial radiologist (ST),blinded to the specific animal treatment, evaluated the μCT images ofall animals to identify and score bony changes and performed all linearmeasurements using Recon software tools.

To quantify the amount of bone loss induced by experimental periodontaldisease (PD), the imaged volume was oriented with the nasal cavity floorparallel to the horizontal plane and the midpalatal suture parallel tothe midsagittal plane. Then the volume was angled such that the longaxis of the distal root of the first molar (D1) and the mesial root ofthe second molar (M2) were vertical to the horizontal plane. Then thedistance between the cementoenamel junction (CEJ) and the alveolar bonecrest (ABC) was measured at the center of D1 and M2. To quantitativelyassess changes in the width of the buccal alveolar outline on axialslices, the imaged volume was oriented such that the floor of the nasalcavity was parallel to the horizontal plane and the midpalatal suturewas parallel to the midsagittal plane. Then the shortest distance fromthe buccal surface of the root to the buccal outline of the alveolarridge was measured for the mesial and distal roots of the first andsecond molars at the level of the hard palate.

The results are summarized in the tables below.

Results

TABLE 1 The effects of ASI on serum parameters CONTROL L L + ASI I L +ASI II P ALP (U/L) 410.29 ± 162.44 323.77 ± 124.98 440.31 ± 69.93 380.54± 85.36  0.061 P (mg/dl) 7.69 ± 1.65 7.98 ± 1.59  8.35 ± 1.82 7.82 ±1.61 0.813 Ca (mg/dl) 9.66 ± 1.17 10.17 ± 0.57  10.37 ± 0.32  9.78 ±0.66^(c) 0.050* CRP (mg/dl) 0.13 ± 0.05  0.26 ± 0.05^(a)   0.16 ±0.07^(b)  0.18 ± 0.06^(b) 0.001** Control: the nonligated treatment. L:the ligature-only; L + ASI I: the ligature plus ASI with dose of 1.81mg/kg of diet. L + ASI II; the ligature plus ASI with dose of 3.62 mg/kgof diet. *p < 0.05, **p < 0.01. ^(a)Significant differences from thegroup as control (p < 0.05). ^(b)Significant differences from the groupas periodontitis (“L”) (p < 0.05). ^(c)Significant differences from thegroup as ASI I (p < 0.05).

TABLE 2 The effects of ASI on several protein levels CONTROL L L + ASI IL + ASI II P IL-1β 100 ± 5.24 189.82 ± 1.84^(a)   142.5 ± 2.53^(a, b) 111.24 ± 2.35^(a, b, c) 0.016* MMP-8 100 ± 2.72 208.72 ± 11.14^(a)166.79 ± 7.86^(a, b) 150.256 ± 11.6^(a, b)  0.019* RANK 100 ± 4.31202.18 ± 2.83^(a)  189.27 ± 5.3^(a, b)  174.31 ± 9.82^(a, b) 0.023*RANKL 100 ± 8.06 167.06 ± 10.72^(a) 150.11 ± 6.34^(a, b) 138.35 ±13.6^(a, b) 0.022* OPG 100 ± 2.3  47.25 ± 2.33^(a)  72.87 ± 4.67^(a, b)  91.13 ± 3.04^(a, b, c) 0.016* MPO 100 ± 6.9  228.86 ± 1.13^(a)    180± 2.97^(a, b)   159.59 ± 13.09^(a, b, c) 0.016* Control: the nonligatedtreatment. L: the ligature-only. L + ASI I: the ligature plus ASI withdose of 1.81 mg/kg of diet. L + ASI II: the ligature plus ASI with doseof 3.62 mg/kg of diet. IL-1β: Interleukin 1β. MMP-8: MatrixMetalloproteinases-8. RANK: Receptor Activator of Nuclear Factor kappaβ. RANKL: Receptor Activator of Nuclear Factor kappa β Ligan. OPG:Osteoprotegerin. MPO: Myeloperoxidase. *p < 0.05. ^(a)Significantdifferences from the group as control (p < 0.05). ^(b)Significantdifferences from the group as periodontitis (“L”) (p < 0.05).^(c)Significant differences from the group as ASI I (p < 0.05)

Table 1 shows that serum CRP levels in ASI treatment groups weresignificantly lower in comparison to the induced periodontitis group.Serum calcium levels were also significantly reduced in the ASI IIgroup.

Table 2 shows that serum levels for IL-1β, MMP-8, RANK, RANKL, OPG, MPOwere significantly lower in comparison to the periodontitis group. Serumlevels for IL-1β and MPO in the ASI II group were significantly lower incomparison to the ASI I group. The results of Table 2 are graphicallydepicted in FIG. 1A.

TABLE 3 Inflammatory cell infiltration (ICI). CONTROL L L + ASI I L +ASI II ICI n (%) n (%) n (%) n (%) P Little 9 (%69.2) 0 (%0)  0 (%0)   0(%0)  0.001** visible Slightly 4 (%30.8)  3 (%23.1) 9 (%69.2) 11 (%84.6)visible Dense 0 (%0)   10 (%76.9) 4 (%30.8)  2 (%15.4) Control: thenonligated treatment. L: the ligature-only. L + ASI I: the ligature plusASI with dose of 1.81 mg/kg of diet. L + ASI II: the ligature plus ASIwith dose of 3.62 mg/kg of diet. **p < 0.01

Table 3 shows that ICI was significantly reduced by ASI treatment. Theresults of Table 3 are graphically depicted in FIG. 2.

FIG. 3 depicts images of alveolar bone (“AK”), ligament (“BD”), dental(“D”), epithelial tissue (“ED”), inflammatory cells (“EH”), pulpa (“P”),periodontal ligament (“PL”). PANEL (A) shows the control: non-ligatedtreatment (HE×40). As shown, there was little visible ICI. PANEL (B)shows the control, non-ligated treatment (HE×100). As shown, there wasno visible ICI. PANEL (C) shows the ligature-only (HE×40). As shown,there was dense ICI. PANEL (D) shows the ligature-only (HE×100). Asshown, there was dense ICI.

PANEL (E) shows the ligature plus ASI with a dose of 1.81 mg/kg of diet(HE×40). As shown, there was less visible ICI. PANEL (F) shows theligature plus ASI with a dose of 1.81 mg/kg of diet (HE×100). As shown,there was less visible ICI. PANEL (G) shows the ligature plus ASI withdose of 3.62 mg/kg of diet (HE×40). As shown, there was less visibleICI. PANEL (H) shows the ligature plus ASI with dose of 3.62 mg/kg ofdiet. (HE×100). As shown, there was less visible ICI.

TABLE 4 Bone Mineral Density (BMD) CONTROL L L + ASI I L + ASI II P BMD0.78 ± 0.03 0.77 ± 0.1 0.78 ± 0.07 0.85 ± 0.06 ^(b) 0.045* (g/cm²)Control: the nonligated treatment L: the ligature-only L + ASI I: theligature plus ASI with dose of 1.81 mg/kg of diet L + ASI II: theligature plus ASI with dose of 3.62 mg/kg of diet. *p < 0.05, ** p <0.01 ^(b) Significant differences from the group as periodontitis.

Table 4 shows that bone mineral density was significantly increased inthe ASI II with respect to the controls and the ASI I group. The resultsof Table 4 are graphically depicted in FIG. 4.

TABLE 5 Mean of bone loss parameters CONTROL L L + ASI I L + ASI II PMCEJ-MCB (mm) 0.91 ± 0.15 (0.88) 1.54 ± 0.14 (1.51) ^(a) 1.35 ± 0.22(1.32) ^(a, b) 1.39 ± 0.12 (1.40) ^(a, b ) 0.001** RF-RFCB (mm) 0.20 ±0.07 (0.20) 0.63 ± 0.11 (0.59) ^(a) 0.46 ± 0.10 (0.44) ^(a, b) 0.53 ±0.12 (0.54) ^(a, b, c) 0.001** DCEJ-DCB (mm) 0.97 ± 0.37 (1.03) 1.29 ±0.24 (1.25)   1.05 ± 0.21 (1.03)    1.26 ± 0.28 (1.26)    0.061 Control:the nonligated treatment L: the ligature-only L + ASI I: the ligatureplus ASI with dose of 1.81 mg/kg of diet; L + ASI II: the ligature plusASI with dose of 3.62 mg/kg of diet. MCEJ: Mesial Cemento-enamelJunction MCB: Mesial Crestal Bone DCEJ: Distal Cemento-enamel JunctionDCB: Distal Crestal Bone RF: Roof of Furcation RFCB: Roof of FurcationCrestal Bone; * p < 0.05, **p < 0.01 ^(a) Significant differences fromthe group as control, (p < 0.001). ^(b) Significant differences from thegroup as periodontitis, (p < 0.001). ^(c) Significant differences fromthe group as ASI I, (p < 0.001).

Table 5 summarizes the mean bone loss results. As shown, mean bone lossbetween the mesial cemento-enamel junction and the mesial crestal bonewas significantly reduced in the treatment groups. Similarly, mean boneloss between the roof of furcation and roof of furcation crestal bonewas significantly reduced in the treatment groups. The results aregraphically depicted in FIG. 5.

TABLE 6 Rate of bone loss in groups CONTROL L L + ASI I L + ASI II PMCEJ- MCB/ 0.28 ± 0.05 0.54 ± 0.04 ^(a) 0.49 ± 0.10 ^(a, b) 0.50 ± 0.10^(a, b) 0.001** MCEJ-MRAP RF-RFCB/ 0.07 ± 0.02 0.26 ± 0.05 ^(a) 0.21 ±0.04 ^(a, b) 0.22 ± 0.05 ^(a, b) 0.001** RF- RFAP DCEJ -DCB/ 0.32 ± 0.120.42 ± 0.09   0.41 ± 0.12    0.42 ± 0.08    0.190 DCEJ- DRAP Control:the nonligated treatment L: the ligature-only L + ASI I: the ligatureplus ASI with dose of 1.81 mg/kg of diet L + ASI II: the ligature plusASI with dose of 3.62 mg/kg of diet. MCEJ: Mesial Cemento-enamelJunction MCB: Mesial Crestal Bone MRAP: Mesial Root Apex DCEJ: DistalCemento-enamel Junction DCB: Distal Crestal Bone DRAP: Distal Root ApexRF: Roof of Furcation RFCB: Roof of Furcation Crestal Bone RFAP: Roof ofFurcation Apex * p < 0.05, **p < 0.01 ^(a) Significant differences fromthe group as control, (p < 0.001). ^(b) Significant differences from thegroup as periodontitis, (p < 0.001).

Table 6 shows the rate of bone loss results. As shown, the rate of boneloss between the mesial cemento-enamel junction and the mesial crestalbone was significantly reduced in the treatment groups. Similarly, therate of bone loss between the roof of furcation and roof of furcationcrestal bone was significantly reduced in the treatment groups. Theresults are graphically depicted in FIG. 6. FIGS. 7-10 depicts 3Dmicro-computed tomography images from the control and treatment groups.As shown, there was less visible tooth and bone damage in the treatmentgroups.

Example 2

The study is a prospective, longitudinal, single-blind pilotintervention trial with six-month follow-up. Subjects presenting withsevere or generalized (at least 50% of teeth affected) periodontitis areinvited to participate in the study. A baseline visit is conducted by ablind calibrated examiner who collects a complete medical history,standard clinical periodontal parameters and blood samples. Thereafter,patients undergo a standard phase of non-surgical periodontal treatmentthat is performed by a periodontist. All other necessary dentaltreatments (extractions of hopeless teeth, restorative treatments) arecarried out prior to completion of the periodontal treatment, consistingof oral hygiene instructions and subgingival scaling and root planing.

The therapeutic phase is completed within 1-3 months of the baselinevisit. Patients are re-examined at 2 and 6 months after the completionof the treatment. The subjects are divided into three groups based onthe baseline visit. Group A includes individuals with marginal (<30%)alveolar bone loss; Group B includes individuals with clinicallysignificant gum recession; and Group C includes individuals withclinically significant enamel erosion. Each group includes 40individuals, who are subdivided into two control groups (C1 and C2) andtwo treatment groups (T1 and T2). The treatment protocol is summarizedin the table below.

TREATMENT GROUP PROTOCOL RESULTS (AT 3 MONTHS) A-T1 Mouthwash twicedaily Significant reduction in alveolar bone loss A-T2 Chewie twicedaily Significant reduction in alveolar bone loss A-C1 Control mouthwashtwice daily No improvement in reducing alveolar bone loss A-C2 Controlchewie twice daily No improvement in reducing alveolar bone loss B-T1Mouthwash twice daily Halt in gum recession B-T2 Chewie twice daily Haltin gum recession B-C1 Control mouthwash twice daily Insignificantimprovement in reducing gum recession B-C2 Control chewie twice dailyInsignificant improvement in reducing gum recession C-T1 Mouthwash twicedaily Halt in enamel erosion C-T2 Chewie twice daily Halt in enamelerosion C-C1 Control mouthwash twice daily Insignificant improvement inreducing enamel erosion C-C2 Control chewie twice daily Insignificantimprovement in reducing enamel erosion

The mouthwash comprises an aqueous solution of arginine silicate andstandard orally acceptable excipients. The chewie comprises abutadiene-based polymer and standard orally acceptable excipientsimpregnated with arginine silicate. The control mouthwash and chewie areidentical to the treatment compositions, but lacking arginine silicate.

Example 3

The study group comprises ten systemically healthy subjects with earlyto mild periodontitis. Patients who have taken antibiotics or receivedperiodontal treatment within 6 months preceding the study are excluded.Prior to any treatment procedure, oral hygiene instructions (OHI) aregiven. Each quadrant of the subjects is randomly assigned to one of thefollowing groups: (A) scaling and root planning combined with dailyadministration of arginine silicate post-treatment, (B) scaling and rootplanning alone, (C) daily administration of arginine silicate alone, and(D) OHI alone.

Subjects are re-examined at one week, four weeks, eight weeks, and 12weeks. At one week, groups (A)-(C) shows similar improvement inperiodontal symptoms, while group (D) shows no significant improvementin periodontal symptoms. At four weeks, Group (A) shows significantimprovement in periodontal symptoms, Groups (B) and (C) show minimalimprovement in periodontal symptoms, and Group (D) shows no improvementin periodontal symptoms. At eight and twelve weeks, Groups (A) and (C)show significant improvement in periodontal symptoms, Group (B) showsminimal improvement in periodontal symptoms, and Group (D) shows noimprovement in periodontal symptoms.

Example 4

20 patients with gingivitis (having at least three of the followingsymptoms: swollen gums, bright red or purple gum tissue, gum tissue thatis sensitive to touch, bleeding gums, bad breath) are studied. Fivepatients are administered once-daily arginine silicate (I); fivepatients are topically administered once-daily arginine silicate to theoral cavity (II); five patients are administered chlorhexidine mouthwashfor twice-daily use (III); and five patients are administered standardover-the-counter mouthwash for twice daily use (IV).

Patients are evaluated at four weeks, eight weeks, and twelve weeks. Atfour weeks, groups (I)-(III) have significant reduction in symptoms ofgingivitis, while group (IV) has only a minimal reduction in symptoms ofgingivitis. At eight and twelve weeks, groups (I) and (II) have the mostsignificant reduction in symptoms of gingivitis, followed by groups(III) and (IV), respectively. The trial is ceased at twelve weeks toavoid side effects from the chlorhexidine treatment. Visible signs ofgingivitis decrease significantly. Gum recession is stopped and/orreversed. Bone mineral density in the oral cavity is also increased.

1.-17. (canceled)
 18. A method of ameliorating gum tissue loss,comprising: identifying an individual having receding gums; andadministering an effective amount of an arginine silicate complex,wherein the amount is effective to stop or reverse gum tissue loss. 19.The method of claim 18, wherein the effective amount of thearginine-silicate complex is topically administered to the oral cavity.20. The method of claim 18, wherein the arginine-silicate complexcomprises arginine, silicate, and inositol.
 21. The method of claim 18,wherein the arginine-silicate complex is formulated as a mouthwash. 22.The method of claim 18, wherein the identifying an individual havingperiodontal disease or symptoms thereof comprises physical examinationby a dental hygienist, dentist, oral surgeon, orthodontist, or medicaldoctor.
 23. The method of claim 18, wherein the effective amount of thearginine-silicate complex is topically administered to the oral cavity.